We have shown that the inducible nitric oxide synthase (iNOS) is expressed in the liver under conditions of sepsis, endotoxemia, and inflammation. In addition, our group has shown that iNOS expression is under the regulation of inflammatory cytokines, and results in synthesis of large amounts of nitric oxide (NO) in both rodent and human hepatocytes. We have also presented evidence that the high but transient expression of iNOS in endotoxemia protects the liver from damage and modulates the activity of specific target enzymes. Others have shown, outside the liver, that sustained iNOS expression occurring as part of chronic inflammatory processes results in tissue dysfunction or damage. Based on these and other observations, we propose a two-part hypothesis to explain the actions of NO in the liver in sepsis. In the first part, we hypothesize that iNOS is expressed transiently in the acute septic response to perform adaptive autocrine and paracrine functions aimed at regulating metabolism and resolving or counteracting the consequences of local immune activation, thus protecting the liver from injury. In the second part, we propose that when iNOS is overexpressed in a sustained manner, as would be seen in septic ICU patients with overwhelming sepsis or the systemic inflammatory response syndrome, NO will have damaging effects in the liver and contribute to organ dysfunction and damage. We will test these hypotheses in carefully selected models and assays in three inter-related aims. AIM I: TO DETERMINE THE AUTOCRINE ACTIONS OF NO ON LIVER METABOLISM IN SEPSIS. We have strong preliminary evidence that iNOS expression in the liver inhibits the activity of glyceraldehyde 3-phosphate dehydrogenase (GAPDH), a gluconeogenic enzyme. Therefore, the autocrine actions of NO on hepatocyte GAPDH activity will be determined for both short-term and long- term iNOS expression. AIM II: TO DETERMINE THE ADAPTIVE PARACRINE ACTIONS OF NO IN THE LIVER IN SEPSIS. Guided by our in vivo data, we will isolate the protective roles of NO in regulating intrahepatic resistance, PMN accumulation, and platelet aggregation in an isolated, perfused liver model. AIM III: TO DETERMINE THE TOXICITY OF SUSTAINED NO SYNTHESIS IN HEPATOCYTES IN SEPSIS. Toxicity of sustained iNOS expression will be determined in in vitro and in vivo sepsis models to determine if NO causes necrosis, heat shock stress response, or apoptosis and/or DNA damage. These experiments will fully test our hypotheses and isolate the actions of NO in the liver in sepsis.